Current status of phylogenetic studies on ciliated protists (Alveolata, Protozoa, Ciliophora) by the OUC-group: advances, challenges and future perspectives

Abstract

Ciliated protists (ciliates) represent a morphologically and genetically diverse group of single-celled eukaryotes, the phylogeny of which is critical for understanding eukaryotic evolution. Through international collaborations, the Laboratory of Protozoology at Ocean University of China (OUC-group) has conducted detailed research on ciliate phylogeny based on expanded taxonomic sampling, employing single gene as well as multi-gene markers, and phylogenomic datasets. We have systematically investigated > 1000 ciliate species spanning ~ 40 orders, sampled from diverse biotopes including marine environments in China seas and freshwater wetlands. This comprehensive sampling has generated three key datasets: (1) genomic DNA extracts from ~ 2600 strains, (2) ~ 2300 sequences of marker genes, and (3) single-cell genomic and/or transcriptomic datasets from ~ 120 species. Based on these datasets, the phylogenetic relationships covering all classes and most orders have been thoroughly reconstructed and investigated, resulting in the establishment of 93 new supraspecies taxa comprising two classes (Mesodiniea and Protocruziea), two subclasses (Protohypotrichia and Synhymenia), two orders (Wilbertomorphida and Lynnellida), 11 families, and 76 genera. Moreover, we have reconstructed a genome-scale tree of life for ciliates and provided an updated classification of the phylum Ciliophora. Furthermore, based on the robust phylogenetic tree of ciliates, we provide more reliable estimates for the origins and divergence times of the main ciliate groups. Future studies integrating advanced genomics, innovations in culturing and interdisciplinary applications will refine the ciliate tree of life, with broader impacts for our understanding of eukaryotic evolution and biodiversity.

Keywords

Ciliates / On-going challenges / Phylogeny and evolution / Phylogenomics / rDNA

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[194] Yi Z, Song W, Clamp JC, Chen Z, Gao S, Zhang Q. Reconsideration of systematic relationships within the order Euplotida (Protista, Ciliophora) using new sequences of the gene coding for small-subunit rRNA and testing the use of combined data sets to construct phylogenies of the Diophrys-complex. Mol Phylogenet Evol, 2009, 50: 599-607

[195] Yi Z, Katz LA, Song W. Assessing whether alpha-tubulin sequences are suitable for phylogenetic reconstruction of Ciliophora with insights into its evolution in euplotids. PLoS ONE, 2012, 7 e40635

[196] Zhang TY, Vďačný P. Deciphering phylogenetic relationships of and delimiting species boundaries within the controversial ciliate genus Conchophthirus using an integrative morpho-evo approach. Mol Phylogenet Evol, 2024, 190 107931

[197] Zhang QQ, Simpson A, Song WB. Insights into the phylogeny of systematically controversial haptorian ciliates (Ciliophora, Litostomatea) based on multigene analyses. P Roy Soc b: Biol Sci, 2012, 279: 2625-2635

[198] Zhang TT, Fan XP, Gao F, Al-Farraj SA, El-Serehy HA, Song WB. Further analyses on the phylogeny of the subclass Scuticociliatia (Protozoa, Ciliophora) based on both nuclear and mitochondrial data. Mol Phylogenet Evol, 2019, 139 106565

[199] Zhang TY, Dong JY, Cheng T, Duan LL, Shao C. Reconsideration on taxonomy of the marine ciliate Neobakuella aenigmatica Moon et al. 2019 (Protozoa, Ciliophora, Hypotrichia). Mar Life Sci Technol, 2020, 2: 97-108

[200] Zhang GAT, Zhao Y, Chi Y, Warren A, Pan HB, Song WB. Updating the phylogeny and taxonomy of pleurostomatid ciliates (Protista: Ciliophora) with establishment of a new family, a new genus and two new species. Zool J Linn Soc, 2022, 196: 105-123

[201] Zhang TT, Tang QY, Chen ZF, Lian CY, Yang XQ, Song WB, Warren A, Shao C. Insights into the phylogeny of the family Deviatidae (Protozoa, Ciliophora, Hypotrichia) based on multi-gene, morphological and ontogenetic information, with the establishment of a new species Deviata multilineae n. sp. Mol Phylogenet Evol, 2022, 177 107623

[202] Zhang TY, Shao C, Zhang TT, Song WB, Vďačný P, Al-Farraj SA, Wang YR (2022c) Multi-gene phylogeny of the ciliate genus Trachelostyla (Ciliophora, Hypotrichia), with integrative description of two species, Trachelostyla multinucleata spec. nov. and T. pediculiformis (Cohn, 1866). Front Microbiol 12:775570

[203] Zhang GAT, Li Y, Gong RT, Qiao Y, Al-Farraj SA, Pan HB, Wang Z, Hines HN. Taxonomy and molecular phylogeny of pleurostomatid ciliates from China with a description of two new species. Protist, 2023, 174 125975

[204] Zhang Y, Dou H, Fu Y, Liang F, Wang Z, Warren A, Li L. Comparative genomics provides insights into the phylogeny and environmental adaptations of Peritrichia (Protista, Ciliophora) – a potential resource for environmental pollution control and bioremediation. Mol Phylogenet Evol, 2023, 186 107835

[205] Zhang GAT, Liu YQ, Pan HB, Liu YJ, Ma HG, Wang Z, Al-Rasheid KAS, Song WB, Hines HN. Diversity of pleurostomatid ciliates: morphology, taxonomy and molecular phylogeny of freshwater isolates found in a northern China wetland, with a description of two new species. Diversity, 2024, 16 294

[206] Zhang GAT, Wang CC, Niu JH, Cao X, Wang Y, Al-Farraj SA, Warren A, Wang Z, Pan HB. Four freshwater cyrtophorids from the Lake Weishan wetland, China (Protozoa, Ciliophora): taxonomy and phylogeny, with description of a new species. Syst Biodivers, 2024, 22 2379860

[207] Zhang GAT, Zhang TY, Chi Y, Pan HB, Zhang Z, Zhang JC, Niu JH, Ma HG, Al-Rasheid KAS, Warren A, Song WB. New insights and investigation: morphomolecular notes on the infraciliature, taxonomy, and systematics of pleurostomatid ciliates (Protozoa, Ciliophora), with establishment of a new suborder, two new genera, and three new species. Zool Res, 2024, 45: 1327-1346

[208] Zhang Y, Fu Y, Vďačný P, Liang F, Dou H, Warren A, Li L. Comparative genomics reveals the evolutionary history of the unicellular eukaryote class Litostomatea and its adaptive evolution based on biochemical metabolic capacity. Zool J Linn Soc, 2024, 201 zlae077

[209] Zhang Y, Liang F, Fu Y, Chi Y, Song W, Warren A, Li L. Comparative genomics provides insights into the evolutionary history of the phylum Ciliophora (Eukaryota, Alveolata) and uncovers the adaptive evolution of anaerobic ciliate classes. Zool Scr, 2024, 54: 270-290

[210] Zhao Y, Yi Z, Warren A, Song WB. Species delimitation for the molecular taxonomy and ecology of the widely distributed microbial eukaryote genus Euplotes (Alveolata, Ciliophora). Proc Biol Sci, 2018, 28520172159

[211] Zheng W, Dou H, Li C, Al-Farraj S, Byerly A, Stover N, Song W, Chen X, Li L. Comparative genome analysis reveals cis-regulatory elements on gene-sized chromosomes of ciliated protists. Front Microbiol, 2022, 13 775646

[212] Zhou Z, Li C, Yuan Q, Chi Y, Li Y, Yan Y, Al- Farraj S, Stover N, Chen Z, Chen X. Single-cell transcriptomic analysis reveals genome evolution in predatory litostomatean ciliates. Eur J Protistol, 2024, 93 126062

[213] Zhuang W, Li R, Feng X, Al-Farraj SA, Hu X. New Contribution to the diversity of the anaerobic genus Metopus (Ciliophora, Armophorea), with descriptions of three new marine species. Front Mar Sci, 2022, 9 884834

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